Single-cell-isolated Trypanosoma cruzi clones were obtained from Triatoma infestans, fed on either a chronic chagasic patient (ten clones) or an animal infected with T. cruzi strain CA-I (nine clones). The growth kinetics in liquid medium (LIT) and peak Coulter volume of these clones were characterized. Significant inter- and intra-group differences in growth rate and peak modal volumes were found. These data indicate that genetically heterogeneous subpopulations of T. cruzi exist in chronic chagasic patients and in T. cruze strains. A simple protocol was developed for serial in vitro cultivation of the intra-cellular amastigote form of T. cruzi in cell-free medium (LIT). The amastigotes are resistant to complement-mediated lysis. In unperturbed cultures, amastigotes grow for two cycles and differentiate to epimastigotes which begin a second exponential growth phase. Continuous passage of growth-phase amastigotes to fresh medium inhibits transformation to epimastigotes.